Consequences of replicative aging in Cryptococcus neoformans Cryptococcus neoformans is a major fungal pathogen that causes a debilitating chronic meningoencephalitis predominantly in immunocompromised patients. This fungal infection is notoriously difficult to treat and despite effective antifungal therapy mortality can be high in patients with progressed disease. Ample of evidence suggests that microevolution of the pathogen population occurs during chronic infection and facilitates persistent disease. Strong support for such a mechanism comes from human, in vitro and animal model data. Although it is evident that microbial factors differ among strains and contribute to difference in virulence these factors are still poorly understood. We determined that C. neoformans undergoes replicative aging and that older cells or more resistant to antifungal treatment when compared to the younger cells. These could potentially give them a biological advantage in vivo and promote their selection under certain conditions. This application proposes to establish if replicative aging of C. neoformans affects the pathogenesis of chronic cryptococcosis. In the first Aim we propose to establish the characteristics that distinguish senescent from young C. neoformans cells. In the second Aim we propose to establish the fate of senescent C. neoformans cells in vivo under antifungal selection pressure. PUBLIC HEALTH RELEVANCE: This grant proposal studies what happens when Cryptococcus neoformans, a fungus that can causes disease in humans grows old. We will investigate how best to distinguish old fungal cells from young fungal cells and if this natural process of aging in this fungus makes this fungus more resistant to therapy. This may facilitate its selection in the host.